COLLABORATION AND SERVICE PROJECT SUMMARY Glycosaminoglycans (GAGs), such as heparin, heparan sulfate (HS), and chondroitin sulfate (CS), are naturally occurring polydisperse linear polysaccharides that are heavily O- and N-sulfated. The interaction between GAGs and proteins are critical for many biological processes including cell-cell and cell-matrix interactions, cell migration and proliferation, growth factor sequestration, chemokine and cytokine activation, microbial recognition and tissue morphogenesis during embryonic development. Hundreds of HS-binding proteins have been identified, but the oligosaccharide structures that mediate particular interactions have been defined in only a few cases due to the structural complexity of HS. The goals of Collaboration and Service in the Resource are to provide an expanded application of our technologies to the scientific community. The Resource has a long and extensive record of collaborations and service with outside investigators. While a number of the collaboration and service projects involve the isolation of GAGs from cells or tissues, compositional disaccharide analysis using SAX-HPLC, HPAEC or ESI-MSn, oligosaccharide mapping by LC- ESI-MS, and structural characterization of individual oligosaccharides by LC, MSn, and NMR, they extend to a broad range of other aspects including synthesis, and cellular and structural biology. The projects can also involve development of MD simulations through GLYCAM web-tools or synthesis of oligosaccharides. A cross section of these is represented in the projects highlighted in the Attachment to this section. Building on our reputation in this area, we will continue to expand on technologies already developed for C&S activities with added capabilities emanating from the TR&D and DBP components in the next funding period, thus enhancing our offerings to the broader scientific community. Within these broader principles, three specific areas in which we will augment our capabilities include: (Aim 1) development of a heparan sulfate array platform using chemical synthesis of well-defined heparan sulfate oligosaccharide structures, (Aim 2) development of GLYCAM online tool enhancements including enhancements in the tools for GAG studies, capabilities for in silico screening of virtual GAG/glycan arrays, in silico conjugation of GAGs to proteins for modeling proteoglycans, and capabilities for short MD simulations of GAG-protein complexes, and (Aim 3) implementation of new analytical tools like MS capabilities including cutting edge technologies in throughput, sensitivity, and our structure determination using the proposed Orbitrap Fusion mass spectrometer. GAG characterization will also be enhanced with new NMR technologies developed at the CCRC using 2- dimensional 1H-15N-HSQC experiments. The 13C optimized cryogenic probe being requested will enhance access to high resolution 13C measurements on GAGs providing improved resolution in the carbon dimension over what can be obtained in the more sensitive 2D HSQC experiments, and also visualize non-protonated carbons.